Nestin is one of the intermediate filaments, together with vimentin and glial fibrillary acidic protein (GFAP), and is detected abundantly in neuroepithelial stem/progenitor cells in the growing central nervous system of embryonal rats and humans (Lendahl, U., et al., Cell (1990) 60: 585-595; Messam, C. A., et al., Exp. Neurol. (2000) 161: 585-596; Tohyama, T., et al., Lab. Invest. (1992) 66: 303-313; Tohyama, T, et al., Am. J. Pathol. (1993) 143: 258-268)). Nestin forms intermediate filament bundles, perhaps with vimentin, by copolymerization in neuroepithelial cells (Eliasson, C., et al., J. Biol. Chem. (1999) 274: 23996-24006; Rutka, J. T., et al., Int. J. Dev. Neurosci. (1999) 17: 503-515).
Nestin mRNA is expressed highly in the cerebrum of developing rat embryos at embryonic day 15 (E15), declines toward postnatal day 12 (P12), and disappears from P18 to the adult stage (Lendahl, U., et al., supra). Using nestin transgene-promoted β-galactosidase expression analysis in mice, LacZ activity has been detected in the neuroepithelium and somites shortly after neural tube closure (E9) (Zimmerman, L., et al., Neuron (1994) 12: 11-24). The LacZ staining becomes stronger in the proliferative ventricular zones of the mouse embryonic striatum and cerebral cortex at E14.5 and E16.5 and decreases in expression in the adult cortex, becoming restricted to a population of ependymal cells.
Substantial nestin expression has also been detected in human gliomas and glioblastomas (Dahlstrand, J., et al., Cancer Res (1992) 52: 5334-5341). Nestin immunostaining has frequently been observed in highly malignant gliomas, especially glioblastomas, as compared with the less malignant forms such as pilocytic astrocytomas. In contrast, nestin is rarely detected by immunostaining in non-neoplastic brain tissues, occurring sometimes faintly in vascular endothelial cells.
Nestin mRNA is approximately 6.2 kilobases long, and its gene contains three introns. Interestingly, neuroepithelium-specific nestin expression is driven by the second intron of the nestin gene, whereas muscle precursor-specific expression is driven by the first intron (Lothian, C., et al., Eur. J. Neurosci. (1997) 9: 452-462; Zimmerman, L, et al., supra).
Nestin expression was previously examined in seven human glioma/glioblastoma-derived culture cell lines (Kurihara, H., et al., Gene Ther. (2000) 7: 686-693). The level of expression varied from high (U251, KG-1C) to non-detectable (NP-2, LN-Z308, T98G) according to Northern blot analysis. The expression levels did not parallel the growth rates of the cell lines, although the degree of malignancy generally reflects tumor doubling time in vivo. The neuronal cell-specific regulator, consisting of the second intron before the 5′ upstream region of the gene, drove LacZ expression in parallel with the extent of mRNA expression in each cell line (Kurihara, H., et al., supra). This variability in nestin expression levels in the glioma/glioblastoma cell lines caused the reevaluation of nestin expression in human glioma/glioblastomas from low to high malignancy grades.
Although a number of angiogenesis-related genes are reported in colorectal cancer endothelium, the nestin gene is not included in the list (Croix, B. S., et al., Science (2000) 289: 1197-1202). Angiogenesis-related genes in brain tumor endothelium may be different from those in colorectal endothelium. It is noteworthy that strong nestin expression is found in brain tumor endothelium even if no nestin expression is found in the brain tumor cells.